Two types of signals are required for T cell activation and proliferation. The first gives specificity to the immune response and involves an interaction between the T-cell receptor/CD3 complex and an antigenic peptide presented by major histocompatibility complex (MHC) class I or class II proteins on the surface of an antigen-presenting cell (APC). The second, called a costimulatory signal, involves interaction between receptor-ligand pairs expressed on the surface of APCs and T cells. Antigenic stimulation in the absence of costimulation induces a state of unresponsiveness or anergy and eventual cell death by apoptosis in the responding T cells. Thus, antigenic stimulation in the presence of costimulation prevents anergy and cell death, thereby promoting cell survival.
A number of investigators have demonstrated that expression of the costimulatory ligands B7-1 and B7-2 on tumor cells can significantly increase immunogenicity of tumors, and induce tumor rejection by a T cell-dependent mechanism. (Baskar, S., Ostrand-Rosenberg, S., Nabavi, N., and Glimcher, L. (1993). Proc. Natl. Acad. Sci. USA. 90, 7015-7019; Chen, L., Ashe, S., Brady, W. A., Hellstrom, I., Hellstrom, K. E., Ledbetter, J. A., McGowan, P., and Linsley, P. S. (1992) Cell 71, 1093-102; Ramarathinam, L., Castle, M., Wu, Y., and Liu, Y. (1994). J Exp Med 179, 1205-14; Townsend, S. E., and Allison, J. P. (1993) Science 259, 368-370). CD28 and CTL4A are two T cell receptors that bind to the B7-1 and B7-2 ligands. CD28 is a transmembrane homodimer that is constitutively expressed on 90% of mammalian CD4+T cells. Engagement of CD28 by its ligands B7-1 or B7-2 on the surface of APCs initiates a signaling cascade culminating in cytokine production and expansion of specific T-cells. CTLA-4, a structural homologue of CD28, is a transmembrane protein that is expressed on activated T cells. The role of B7-CTLA4 interaction remains controversial. (Liu, Y. (1997). Immunol Today 18, 569-72).
Anti-CTLA4 Antibodies
Allison and colleagues have demonstrated that anti-mouse CTLA4 mAb induced rejection of tumors in vivo (Leach et al., 1996) Anti-CTLA4 mAb-treatment has also been shown to enhance immune response of mouse to bacteria. (Heinzel, F. P., and Maier, R. A., Jr. (1999). Infect Immun 67, 6454-60; Kirman, J., McCoy, K., Hook, S., Prout, M., Delahunt, B., Orme, I., Frank, A., and Le Gros, G. (1999). Infect Immun 67, 3786-92).
CTLA4 has also been shown to play an important role in autoimmune diseases. Numerous studies have linked the polymorphism of the CTLA4 gene in humans to a variety of autoimmune diseases, such as Graves' disease, Hashimoto's thyroiditis, myasthenia gravis with thymoma, and insulin-dependent diabetes mellitus. (Barbesino et al., 1998; Braun et al., 1998; Donner et al., 1997; Donner et al., 1997; Huang et al., 2000; Huang et al., 1998; Kotsa et al., 1997; Marron et al., 1997; Nistico et al., 1996; Tomer et al., 1997). In addition, treatment with anti-CTLA4 mAbs has been shown to exacerbate experimental autoimmune diseases such as diabetes (Luhder et al., 1998) and experimental autoimmune encephalomyelitis (animal model of human multiple sclerosis) (Karandikar et al., 1996) in animal models.
In most experimental models, the two most extensively used anti-murine CTLA4 mAbs enhanced T cell activation in vivo. Nevertheless, since different mAbs may react with different epitopes on the same region of a protein or alternatively, bind to the same epitope with different affinities, different mAbs against the same protein can have opposite effects. Thus, some monocloanal antibodies to the CTLA4 receptor may enhance T cell proliferation and activation; while other anti-CTLA4 monoclonal antibodies may inhibit T cell proliferation and activation. Accordingly, monoclonal antibodies and other agents which specifically or selectively interact with CTLA4 may be of therapeutic value in cancer, and infection, as well as autoimmune disease and transplantation.
A major impediment to the development or identification of mAbs or other agents that can be used to either activate or inactivate T cells via a CTLA4-mediated pathway is the lack of an appropriate assays for screening such agents. Conflicting results have demonstrated that an in vitro assay cannot serve as an accurate predictor of the functionality of antibodies to the B7 receptors in vivo. For example, anti-CD28 mAb have been shown to provide potent costimulatory activity for T cell activation in vitro (Jenkins et al., 1991), but have little effect on tumor growth in vivo (Leach et al., 1996). In contrast, investigators have found that administration of anti-CTLA4 mAb induces rejection of tumors and exacerbates autoimmune diseases in vivo (Leach et al.,) while anti-CTLA4 mAbs have been shown to either enhance or inhibit T cell activation in vitro (Krummel and Allison, 1995; Walunas et al., 1994) These results demonstrate that the functional effect of agents targeting receptors for B7 ligands, particularly the CTLA4 receptor, cannot be accurately tested in vitro. Accordingly, it is desirable to have additional tools for screening agents, particularly monoclonal antibodies, that either inhibit or enhance CTLA-4 signaling.